The field of the invention is cell proliferation.
The nematode Caenorhabditis elegans (C. elegans) is well suited for developmental genetic studies because the entire cell lineage has been mapped and is essentially invariant from one animal to the next. Thus, by comparing the cell lineage of a wild-type animal to that of a mutant animal, the changes in cell fates caused by the mutation can be determined.
A number of mutations that alter cell lineage in C. elegans, termed lin mutations, were obtained in genetic screens conducted by Horvitz and Sulston in the late 1970's. A subset of the mutations affected the formation of the vulva, a structure on the ventral surface of C. elegans hermaphrodites through which eggs are laid and through which sperm enters during cross-fertilization. Six vulval precursor cells have the potential to undertake a vulval cell lineage, as defined by the number and pattern of cell divisions. In a wild-type animal only three of these cells actually undertake vulval cell fates and these three cells generate the 22 cells that make up the adult vulva. In multivulva (Muv) animals, most or all of the six vulval precursor cells undertake vulval cell fates. In addition to the cells required for the formation of a normal vulva, these mutant animals generate an excess of cells which cause the formation of raised, vulva-like structures on the ventral surface of the animal. On the other hand, a vulvaless (Vul) phenotype results when no or too few vulval precursor cells adopt vulval cell fates.
Genetic and molecular analyses of Muv and Vul animals have defined a Ras signal transduction pathway that mediates induction of the hermaphrodite vulva. This pathway includes the LIN-3 EGF-like ligand, the LET-23 receptor tyrosine kinase, the SEM-5 adaptor, LET60 Ras, the KSR-1 kinase, LIN-45 MEK-2, and the MPK-1 MAP kinase, and regulates the activities of the ETS transcription factor LIN-1 and the winged-helix transcription factor LIN-31 (reviewed by Horvitz and Sternberg, Nature 351:535-41, 1991; Sundaram and Han, Bioessays 18:473-480, 1996; Tan et al., Cell 93:569-580, 1998). Mutant animals in which this pathway is ectopically activated can display a Muv phenotype, whereas mutant animals that have reduced Ras pathway signaling can display a Vul phenotype.
The synthetic multivalva (synMuv) genes act in two functionally-redundant pathways as negative regulators of the nematode Ras signaling pathway. The first synthetic multivulva mutant was identified by Horvitz and Sulston. The two genetic loci mutated in this mutant were termed lin-8 and lin-9. Reduction-of-function mutations in both of these loci were required for a multivulva phenotype. Subsequent genetic screens identified a set of loci which fall into the same class as lin-8, termed class A genes, and genes which fall into the same class as lin-9, termed class B genes. In general, an animal with a reduction-of-function mutation in any class A gene and a reduction-of-function mutation in any class B gene will display a multivulva phenotype, while animals carrying one or more mutations of the same class have a wild-type vulval phenotype. These two classes appear to define two functionally redundant pathways that negatively regulate the expression of vulval cell fates.
Thus far at least four class A loci (lin-8, lin-15A, lin-38, and lin-56) and at least fourteen class B loci (lin-9, lin-15B, lin-35, lin-36, lin-37, lin-13, lin-52, lin-53, lin-54, lin-55 (dpl-1), lin-61, hda-1, tam-1 (Hsieh et al., Genes & Dev. 13:2958-2970, 1999), and the C. elegans E2F1 homolog (efl-1)) have been identified genetically. lin-15 encodes both A and B activities in two non-overlapping transcripts. In addition, lin-37, lin-35, lin-53, lin-52, lin-54, lin-55 (dpl-1), lin-15A, lin-15B, lin-36, lin-9, lin-55, and efl-1 have been cloned (Ceol and Horvitz, Molecular Cell 7:461-473, 2001; Clark et al., Genetics 137:987-997, 1994; Huang et al., Mol. Biol. Cell 5:395-411, 1994; Beitel et al., Gene 254:253-263, 2000; and PCT WO 98/54299).
A number of the synMuv family members encode polypeptides with sequence similarity to polypeptides involved in cancer development and progression. For example, lin-35 encodes a homolog of the mammalian pocket protein family, which includes retinoblastoma protein (Rb), p107, and p130. This family of proteins has been the subject of intense study since the cloning of Rb in 1986. Rb is a tumor suppressor gene; mutations that inactivate Rb predispose individuals to tumor formation. Most commonly, inactivation of Rb results in a type of eye cancer, retinoblastoma, although inactivating mutations in Rb have been found in other types of tumors. The Rb protein is thought to function as a negative regulator of cell cycle progression. A number of molecules that interact, both directly and indirectly, with Rb and the other pocket proteins have been characterized in mammalian cells.
Another synMuv family member, lin-53, encodes a homolog of p48, a protein which has been shown to bind Rb. Although the functional significance of the interaction between p48 and Rb is not fully understood, recent studies suggest that p48 may play a role in remodeling chromatin structure. In addition, lin-55(dpl-1) encodes a homolog of the DP family of proteins (Ceol and Horvitz, Molecular Cell 7:461-473, 2001). DP family members, together with E2F proteins, bind DNA at specific sites, thereby regulating the transcription of genes that are essential for cell cycle progression. Furthermore, pocket proteins such as Rb bind to the DP-E2F complex to repress transcription.
As in the nematode, Ras pathways have been found to control cell proliferation in a range of organisms from the yeast Saccharomyces cerevisiae to humans. The Ras pathway defines one class of oncogene signaling pathways; members of this pathway, most commonly Ras itself, have been shown to be mutated in a broad range of human cancers (Hunter, Cell 88:333-346, 1997). Accordingly, analysis of the Ras pathway, in particular the vulval induction pathway, in C. elegans addresses the significant need of increasing our understanding of cancer in general.